PROJECT SUMMARY/ABSTRACT Zika virus (ZIKV), a mosquito-borne flavivirus genetically related to dengue virus (DENV), produces a mild febrile illness with maculo-papular rash with approximately 80% of patients demonstrating no symptoms. Until 2007 only 14 cases of human disease had been reported since ZIKV's discovery in 1947. In 2015, an epidemic exploded with estimations of over one million current infections in Brazil alone. While ZIKV infections are generally mild, recently they have been associated with Guillain-Barr syndrome in adults and children and an alarmingly high rate of microcephaly in newborns. There are no commercial vaccines or diagnostic assays available. Accurate detection of ZIKV immunity, as measured by anti-ZIKV antibodies, will be a critical tool in prevention and control programs. Current ZIKV sero-diagnostic methods typically show substantial cross- reactivity with other flavivirus reagents. We have recently developed a revolutionary approach to discover novel diagnostic antibody ligands for viral infections (HIV and DENV) by screening a very large combinatorial library (108 molecular variants) of biologically inspired, but non-biological, molecular shapes, or peptoids, against panels of post-infection patient sera. Using this method, we isolated several peptoids that specifically bound antibodies present in post-infection sera that are not present in normal non-immune sera nor pre- infection sera. Our proof-of-concept preliminary studies have demonstrated the ability of peptoid biomarkers to differentiate between HIV antibody positive and HIV antibody negative serum as well as DEN antibody positive and DEN antibody negative serum. Additionally, we have also identified peptoid biomarkers that can distinguish between recent and non-recent HIV infections. We have used these novel non-biological specific ligands as the bases for HIV ELISA assays which display very good sensitivity and specificity. In this grant, we propose to harness this novel technology to develop robust, peptoid-based assays to specifically detect ZIKV antibodies. We will test this innovative technology against panels of sera, assembled from patients in Recife, the epicenter of the Zika outbreak in Brazil. Direct access to clinical cohorts and patient samples positions our team well for a ZIKV diagnostic development. After identification of ZIKV reactive peptoids, we will perform a detailed characterization of the kinetics of the immunoglobulin responses against the selected peptoids. The identification of these peptoid biomarkers will then lead to the development of a second-generation combinatorial library where peptoid reactivity will be further optimized, and we will identify the mechanisms of binding between the ZIKV-specific peptoid biomarkers and the patient antibody. The goals described in this project will lead to the development of a highly optimized, low cost, peptoid-based high-throughput assay to accurately detect ZIKV antibodies and thereby diagnose individual patients and measure incidence in affected populations. The development of such diagnostic assay and disease surveillance tools will greatly increase the capacity of public health agencies worldwide to monitor infectious diseases of global importance.